Association with actin mediates the EGTA-resistant binding of cytosolic phospholipase A2-alpha to the plasma membrane of activated platelets.
The association of cytosolic phospholipase A(2)-alpha (cPLA(2)alpha) with intracellular membranes is central to the generation of free arachidonic acid and thromboxane A(2) in activated platelets. Despite this, the site and nature of this membrane association has not been fully characterised upon platelet activation. High resolution imaging showed that cPLA(2)alpha was distributed in a partly structured manner throughout the resting platelet. Upon glass activation or thrombin stimulation, cPLA(2)alpha relocated to a peripheral region corresponding to the platelet plasma membrane. Upon thrombin stimulation of platelets a major pool of cPLA(2)alpha was associated with the plasma membrane in an EGTA-resistant manner. EGTA-resistant membrane binding was abolished upon de-polymerisation of actin filaments by DNase I and furthermore, cPLA(2)alpha co-immunoprecipitated with actin upon thrombin stimulation of platelets. Immunofluorescence microscopy studies revealed that, upon platelet activation, cPLA(2)alpha and actin co-localised at the plasma membrane. Thus we have identified a novel mechanism for the interaction of cPLA(2)alpha with its membrane substrate via interaction with actin.
Hastings
Ann D
AD
Faculty of Biological Sciences, University of Leeds, Leeds LS2 9JT, UK.
Herbert
Shane P
SP
Gawler
Debra
D
Walker
John H
JH
D008841:Actin Cytoskeleton / Q000648:ultrastructure; D000199:Actins / Q000378:metabolism* / Q000648:ultrastructure; D016718:Arachidonic Acid / Q000378:metabolism; D001792:Blood Platelets / Q000201:enzymology*; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology*; D003850:Deoxyribonuclease I / Q000378:metabolism; D004533:Egtazic Acid / Q000494:pharmacology*; D054513:Group IV Phospholipases A2 / Q000378:metabolism*; D006801:Humans; D047468:Immunoprecipitation; D008856:Microscopy, Fluorescence; D015539:Platelet Activation / Q000187:drug effects; D021381:Protein Transport; D013917:Thrombin / Q000494:pharmacology; D013928:Thromboxane A2 / Q000494:pharmacology; D013997:Time Factors
D000199:Actins; D016718:Arachidonic Acid; D004533:Egtazic Acid; D013928:Thromboxane A2; D054513:Group IV Phospholipases A2; D003850:Deoxyribonuclease I; D013917:Thrombin